It is our purpose to continue the study of the molecular biology of the human dependovirus, adeno-associated virus (AAV) 2. In the past several years, we have come to believe the AAV is not defective but rather is a virus which preferentially in healthy cells inhibits its own gene expression and DNA replication to enhance the ability of the viral DNA to integrate into the cellular genome. When the cell is infected by adeno- or herpes virus or exposed to toxic treatment, the intracellular milieu is changed to one that is permissive for production of AAV virions. This proposal contains an experimental program designed to investigate the following areas: 1. All of the trans-acting regulatory functions of AAV are encoded in the rep gene. We shall study a) the cis active negative regulation of rep during productive infection; b) negative regulation of AAV promoters by rep under nonpermissive conditions; c) regulation by rep of gene expression at the translational level; d) the mechanism whereby the SV40 regulatory region is controlled by being put into the AAV genome. 2. We shall study the mechanism of replication of cloned AAV genomes with partially deleted terminal repeats and try to establish an in vitro assay for DNA replication. 3. We have preliminary data that suggest that AAV integration in human cells is into a specific sequence on chromosome 21. We wish to confirm this and determine and map the sequence on chromosome 21.